Topical anti-inflammatory pharmaceutical composition with zileuton cream formulation

ABSTRACT

Provided herein is a zileuton cream type topical anti-inflammatory pharmaceutical composition, and more particularly, a zileuton cream type topical anti-inflammatory pharmaceutical composition capable of retaining stability at room temperature and of being applied topically to maximize medical effects while minimizing absorption to an entirety of body, thereby minimizing toxicity caused by the compound so as to be suitable to topical treatment of skin diseases caused by leukotriene.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation-in-part of U.S. patent applicationSer. No. 14/777,183, filed on Sep. 15, 2015, which is a National Stageapplication of International Application Number PCT/KR2014/007525, filedAug. 13, 2014; which claims priority to Korean Patent Application No.10-2013-0130165, filed Oct. 30, 2013; all of which are incorporatedherein by reference in their entirety.

FIELD OF THE INVENTION

The present invention relate to a topical pharmaceutical compositionwith zileuton cream formulation for treatment of skin diseases caused byleukotriene.

BACKGROUND

Leukotriene is a metabolite of arachidonic acid, and is well known as aninflammatory factor that is deeply involved in causing inflammation,edema, and secretion of mucus and the like. A well-known leukotrieneinhibitor developed so far is zileuton ((±)-1-(1-(benzo[b]thiophen-2-yl)ethyl)-1-hydroxyurea).

Zileuton is a mixture of stereoisomers:(R)-1-(1-(benzo[b]thiopen-2-yl)ethyl)-1-hydroxyure and(S)-1-(1-(benzo[b]thiophen-2-yl)ethyl)-1-hydroxyurea. It has beenapproved as an oral drug for treatment of asthma, and is known to beadministrable twice a day. Clinical trials have shown that zileutonexhibits an effective anti-inflammatory pharmaceutical effect againstinflammation reaction of asthma.

Clinical trials have revealed that oral administration of zileutonalleviates atopic dermatitis symptoms. However, zileuton is not beingwidely used despite its excellent anti-inflammatory effects due totoxicity of the compound.

Zileuton has significant hepatotoxicity, and thus an oral administrationof zileuton should be accompanied by monitoring of liver functions ofthe patient.

BRIEF SUMMARY

Therefore, a purpose of the present invention is to provide a topicalpharmaceutical composition with a cream formulation, capable of applyingzileuton topically to skin lesions of diseases caused by topical skininflammation reactions such as atopic dermatitis, acne, various types ofurticaria, psoriasis, eczema, bullous skin diseases such as bullouspemphigoid, collagenoses, Sjogren-Larsson syndrome, mastocytosis and thelike, and maximizing pharmaceutical effects of zileuton while minimizingabsorption to an entirety of a patient's body so as to minimize toxicityof the compound.

Another purpose of the present invention is to provide a cream typetopical pharmaceutical composition having improved physiochemicalstability.

Further, another purpose of the present invention is to provide a methodfor preparing a topical pharmaceutical composition with zileuton creamformulation.

Further, still another purpose of the present invention is to provide amethod for relieving or treating atopic dermatitis, acne, urticaria,psoriasis, eczema, a bullous skin disease, collagenoses, Sjogren-Larssonsyndrome, or acne in skin lesions of mastocytosis using thepharmaceutical composition.

In order to achieve the purposes, one embodiment of the presentinvention provides a cream type topical pharmaceutical composition,comprising zileuton, an aqueous solvent, an organic solvent, athickening agent, and an emulsifier.

The aqueous solvent may comprise water or a pH buffer solution, and theorganic solvent may comprise, without any limitation, at least oneselected from the group consisting of ethanol, polyethylene glycol,hexylene glycol, propylene glycol monocaprylate, benzyl alcohol,N-methyl pyrrolidone, triacetin, isopropyl myristate, propylene glycoldicaprylocaprate, medium chain triglycerides, castor oil, olive oil,safflower oil, light mineral oil, squalane, dimethicone, cyclomethicone,polyglyceryl-6 dioleate, polyglyceryl-10 monooleate, polyglyceryl-10dioleate, polyoxyl 40 hydrogenated castor oil, caprylocaproyl polyoxyl-8glycerides, polyoxyethylene (20) sorbitan monolaurate, diethylene glycolmonoethyl ether, and dimethyl sulfoxide.

The aqueous solvent may be a pH buffer solution having a pH of about 4to about 5.

The thickening agent may comprise at least one of white petrolatum andwhite wax.

The emulsifier may comprise at least one selected from a groupconsisting of phospholipid, stearyl alcohol and propylene glycolstearate. Preferably, the emulsifier may comprise phospholipidsincluding hydrogenated lecithin.

Furthermore, the pharmaceutical composition of the present invention mayfurther comprise a preservative.

The preservative may comprise at least one selected from a groupconsisting of methylparaben, ethylparaben, propylparaben,isobutylparaben, butyl paraben, 2-phenoxy ethanol, and 4-hydroxybenzoicacid.

The pharmaceutical composition of the present invention may comprisezileuton of about 0.05 to about 2 weight % as an active pharmaceuticalingredient; water or pH buffer solution of about 30 to about 34 weight%; white petrolatum of about 36 to about 40 weight %; white wax of about4 to about 8 weight %; propylene glycol of about 16 to about 22 weight%; phospholipid of about 2 to about 6 weight %; and preservative ofabout 0.005 to about 0.04 weight %.

Further, the composition may comprise zileuton of about 0.1 to about 1weight %; water or pH buffer solution of about 31 to about 33 weight %;white petrolatum of about 37 to about 39 weight %; white wax of about 5to about 7 weight %; propylene glycol of about 18 to about 20 weight %;phospholipid of about 3 to about 5 weight %; and preservative of about0.01 to about 0.03 weight %.

The composition may be for topical application to human skin, and thezileuton may be racemic zileuton.

Another embodiment of the present invention provides a method forrelieving or treating atopic dermatitis, acne, urticaria, psoriasis,eczema, a bullous skin disease, collagenoses, Sjogren-Larsson syndrome,or acne in skin lesions of mastocytosis, comprising administering, to asubject in need of such relief or treatment, the composition of thepresent invention.

Still another embodiment of the present invention provides a method forpreparing a cream type topical anti-inflammatory pharmaceuticalcomposition, including mixing zileuton, an aqueous solvent, an organicsolvent, a thickening agent, an emulsifier, and optionally apreservative under a mixing condition of a temperature of about 30 toabout 80° C. and about 600 to about 1200 rpm, for about 10 to about 60minutes.

In the method, the aqueous solvent may be water or pH buffer solutionhaving a pH of about 4 to about 5.

The method may further include cooling the mixed ingredients at atemperature of about 20 and about 30° C. while stirring at about 600 rpmor less.

As aforementioned, the present invention provide a cream type topicalpharmaceutical composition that comprises zileuton as an activepharmaceutical ingredient and exhibits treatment effects to diseasescaused by topical skin inflammation reactions such as atopic dermatitis,acne, various types of urticaria, psoriasis, eczema, bullous skindiseases such as bullous pemphigoid, collagenoses, Sjogren-Larssonsyndrome, skin lesions of mastocytosis, and the like.

Furthermore, the cream type pharmaceutical composition according to thepresent invention has a pharmacokinetic profile that enables zileutonthat is a leukotriene inhibitor to be absorbed to skin effectively butminimizing the amount of absorption to an entirety of a patient's body,and a physiochemical stability.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph showing the pH changes of the pharmaceuticalcompositions with or without a pH buffer solution at 40° C. over 4weeks.

FIG. 2 is a schematic diagram illustrating results of a test conductedto verify medical effects of the pharmaceutical composition according tothe present invention by applying the same to an atopic mouse model;

FIG. 3 is a graph showing thicknesses of a mouse's ear measured aftertopically applying the pharmaceutical composition according to thepresent invention to an atopic mouse model; and

FIGS. 4 and 5 show research results of the pharmacokinetic profile ofthe pharmaceutical composition according to the present invention in aminipig.

DETAILED DESCRIPTION

The present invention now will be described more fully hereinafter inthe following detailed description of the invention, in which some, butnot all embodiments of the invention are described. Indeed, the presentinvention may be embodied in many different forms and should not beconstrued as limited to the embodiments set forth herein; rather, theseembodiments are provided so that this disclosure will satisfy applicablelegal requirements.

The following detailed description is provided to assist the reader ingaining a comprehensive understanding of the compositions and methodsdescribed herein. Accordingly, various changes, modifications, andequivalents of the compositions and/or methods described herein will besuggested to those of ordinary skill in the art. Also, descriptions ofwell-known functions and constructions may be omitted for increasedclarity and conciseness.

Furthermore, a singular form may include a plural from as long as it isnot specifically mentioned in a sentence. Furthermore,“comprise/includes” or “comprising/including” used in the specificationrepresents that one or more components, steps, operations, and elementsexist or are added.

Furthermore, unless defined otherwise, all the terms used in thisspecification including technical and scientific terms have the samemeanings as would be generally understood by those skilled in therelated art. The terms defined in generally used dictionaries should beconstrued as having the same meanings as would be construed in thecontext of the related art, and unless clearly defined otherwise in thisspecification, should not be construed as having idealistic or overlyformal meanings.

Herein below, explanation will be made on a pharmaceutical compositionaccording to the present invention, a method for preparing the same, anda use thereof.

In order to provide a cream type topical formulation comprising zileutonas an active pharmaceutical ingredient, which exhibits effectivetreatment effects to atopic dermatitis and acne and the like, theinventors of the present invention sought for ingredients that can beapplied with zileuton among the ingredients being used in cream typeformulations, and sought for amount ratio of the ingredients forpreparing a cream formulation comprising them, and then defined optimalconcentration of the zileuton and stabilization conditions for realizingphysiochemical stability, and thereby completed the present invention.

Therefore, the present invention provides a cream type topicalpharmaceutical composition comprising zileuton, an aqueous solvent, anorganic solvent, a thickening agent and an emulsifier.

Herein, zileuton is a mixture of stereoisomer:(R)-1-(1-(benzo[b]thiopen-2-yl)ethyl)-1-hydroxyure and(S)-1-(1-(benzo[b]thiophen-2-yl)ethyl)-1-hydroxyurea, in a white powderform, and may be racemic zileuton. That is, the present inventionprovides a cream type composition optimized to racemic zileuton as anactive pharmaceutical ingredient.

The zileuton may be synthesized chemically, or is available commercially(Cornerstone Therapeutics Inc., product name: ZYFLO, ZYFLO CR etc.)

The aqueous solvent used in the pharmaceutical composition of thepresent invention may comprise water or a pH buffer solution.

In a preferred embodiment of the present invention, the aqueous solventis a pH buffer solution. In a more preferred embodiment, the aqueoussolvent may be a pH buffer solution having a pH of about 4 to about 5.For example, the pH buffer solution may be, but not limited to, citratebuffer (citric acid with sodium citrate), acetate buffer (acetic acidwith sodium acetate) or phthalate buffer (potassium hydrogen phthalatewith sodium hydroxide or formaldehyde).

In case of using such pH buffer solution as an aqueous solution, thepharmaceutical composition of the present invention may exhibit moreimproved chemical stability. This is because the pH buffer solutioncontrols the pH change of the composition so that any possiblehydrolysis of zileuton to produce hydrolytic degradants which areundesirable impurities in the composition can be prevented, and therebythe chemical stability of the composition can be improved.

In a preferred embodiment of the present invention, the production ofhydrolytic degradants of zileuton can be minimized by stabilizing the pHof the composition by using a pH buffer solution, and thereby thechemical stability of the composition may be improved.

The organic solvent used in the pharmaceutical composition of thepresent invention may comprise, but not limited to, at least oneselected from the group consisting of ethanol, polyethylene glycol,hexylene glycol, propylene glycol monocaprylate, benzyl alcohol,N-methyl pyrrolidone, triacetin, isopropyl myristate, propylene glycoldicaprylocaprate, medium chain triglycerides, castor oil, olive oil,safflower oil, light mineral oil, squalane, dimethicone, cyclomethicone,polyglyceryl-6 dioleate, polyglyceryl-10 monooleate, polyglyceryl-10dioleate, polyoxyl 40 hydrogenated castor oil, caprylocaproyl polyoxyl-8glycerides, polyoxyethylene (20) sorbitan monolaurate, diethylene glycolmonoethyl ether, and dimethyl sulfoxide.

In a preferred embodiment of the present invention, the organic solventis propylene glycol.

The thickening agent used in the pharmaceutical composition of thepresent invention may comprise, but not limited to, at least one ofwhite petrolatum and white wax.

The emulsifier may be, but not limited to, at least one selected from agroup consisting of hydrogenated lecithin, stearyl alcohol and propyleneglycol stearate. Preferably, the emulsifier may comprise phospholipidsincluding hydrogenated lecithin. For example, the emulsifier iscommercially available as a product name of Phospholipon 90H.

The pharmaceutical composition of the present invention may furthercomprise a preservative.

The preservative may be, but not limited to, at least one selected froma group consisting of methylparaben, ethylparaben, propylparaben,isobutylparaben, butyl paraben, 2-phenoxy ethanol, and 4-hydroxybenzoicacid.

The pH buffer solution, organic solvent, thickening agent such as whitepetrolatum and white wax, emulsifier, and preservative to be used in thepharmaceutical composition of the present invention are availablecommercially.

In one exemplary embodiment, the cream type pharmaceutical compositionof the present invention may comprise zileuton of about 0.05 to about 2weight % as an active pharmaceutical ingredient; water or pH buffersolution of about 30 to about 34 weight %; white petrolatum of about 36to about 40 weight %; white wax of about 4 to about 8 weight %;propylene glycol of about 16 to about 22 weight %; phospholipid of about2 to about 6 weight %; and preservative of about 0.005 to about 0.04weight %.

In another exemplary embodiment, the cream type pharmaceuticalcomposition of the present invention may comprise zileuton of about 0.1to about 1 weight %; water or pH buffer solution of about 31 to about 33weight %; white petrolatum of about 37 to about 39 weight %; white waxof about 5 to about 7 weight %; propylene glycol of about 18 to about 20weight %; phospholipid of about 3 to about 5 weight %; and preservativeof about 0.01 to about 0.03 weight %.

The cream type topical pharmaceutical composition of the presentinvention may retain its physical and chemical stability for about 4weeks or more under the temperature condition of about 25 to about 40°C.

The present invention also provides a method for relieving or treatingatopic dermatitis, acne, urticaria, psoriasis, eczema, a bullous skindisease, collagenoses, Sjogren-Larsson syndrome, or acne in skin lesionsof mastocytosis, wherein said method comprises administering, to asubject in need of such relief or treatment, a composition of thepresent invention.

In the method for relieving or treating any of the aforementioneddiseases according to the present invention, the composition may beapplied topically to human skin.

According to an embodiment of the present invention, the medical effectof the cream type topical pharmaceutical composition has been verifiedin a Delayed Type Hypersensitivity Reaction in mouse induced by DNFB.The test article was applied to a mouse's skin 3 times a day, and thenthe medical effect was measured. Furthermore, an excess amount of thezileuton was injected using a non-optimized test vehicle, acetone. Testresults showed that an increase of thickness of the mouse's ear causedby inflammation and edema in the mouse model administered with the creamtype pharmaceutical composition of the present invention was effectivelyinhibited, proving that the composition has an effectiveanti-inflammation function.

In order to determine whether or not a medical effect caused byabsorption to an entirety of body is included, absorption to skin andbody absorption pattern were evaluated. For this purpose, a skin PK wasexamined from a minipig having a skin structure most similar to human.Test results showed effective skin absorption patterns, but showed nosystemic exposure pattern. That is, it was identified that a cream typetopical pharmaceutical composition according to the present inventioneffectively induces skin absorption, and minimizes body absorption,proving that the composition according to the present invention issuitable formulation for administration to human skin.

Therefore, a cream type topical pharmaceutical composition according tothe present invention exhibits an anti-inflammatory effect by inhibitingformation of leukotriene, and thus is effective in relieving or treatingdiseases caused by topical skin inflammation reaction such as atopicdermatitis, acne, various types of urticaria, psoriasis, eczema, bullousskin diseases such as bullous pemphigoid, collagenoses, Sjogren-Larssonsyndrome, skin lesions of mastocytosis, and the like.

The acne may be an inflammatory acne selected from a group consisting ofacne papulosa, acne pustulosa, acne papulopustolosa, and severeinflammatory acne.

The cream type pharmaceutical composition may be for topical applicationto human skin. More specifically, it may be applied topically once tofour times a day.

Further, the present invention also provides a preparation method for acream type topical pharmaceutical composition, the method comprisingmixing zileuton, an aqueous solvent, an organic solvent, a thickeningagent, and emulsifier under a mixing condition of a temperature of 30and 80° C. and 600 to 1200 rpm.

The mixing may be performed for 10 to 60 minutes.

The aqueous solvent used in the method may comprise a pH buffersolution. In a preferred embodiment, the aqueous solvent may be, a pHbuffer solution having a pH of about 4 to about 5. For example, the pHbuffer solution may be, but not limited to, citrate buffer (citric acidwith sodium citrate), acetate buffer (acetic acid with sodium acetate)or phthalate buffer (potassium hydrogen phthalate with sodium hydroxideor formaldehyde).

In the mixing step, a preservative may be further mixed.

In one exemplary embodiment, the preparation method for a cream typetopical pharmaceutical composition of the present invention may comprisemixing zileuton of about 0.05 to about 2 weight %; water or pH buffersolution of about 30 to about 34 weight %; white petrolatum of about 36to about 40 weight %; white wax of about 4 to about 8 weight %;propylene glycol of about 16 to about 22 weight %; phospholipid of about2 to about 6 weight %; and preservative of about 0.005 to about 0.04weight %.

In another exemplary embodiment, the preparation method for a cream typetopical pharmaceutical composition of the present invention may comprisemixing zileuton of about 0.1 to about 1 weight %; water or pH buffersolution of about 31 to about 33 weight %; white petrolatum of about 37to about 39 weight %; white wax of about 5 to about 7 weight %;propylene glycol of about 18 to about 20 weight %; phospholipid of about3 to about 5 weight %; and preservative of about 0.01 to about 0.03weight %.

The preparation method for a cream type topical pharmaceuticalcomposition may further comprise cooling the resultant from the mixingunder the condition of a temperature of about 20 and about 30° C. whilestirring at about 600 rpm or less.

The cream type topical pharmaceutical composition prepared through theaforementioned method may retain its stability for about 4 weeks or moreat a temperature of about 25 to about 40° C., and may thus be suitableto topical application.

Hereinafter, examples of a cream type topical pharmaceutical compositionof the present invention will be explained. However, the examples areintended for just illustrating, but not limiting the scope of thepresent invention thereto.

Example 1 Preparing Zileuton Cream Type Composition

1. Screening Test for Suitability of the Cream Type Composition

Suitability of components generally used in cream formulation withrespect to zieluton was screened, prior to preparing a zieluton creamformulation. For this purpose, 5 mg of zileuton was dissolved in 5 mg ofexcipient, and then changes of TRS % (Total Related Substances) wereobserved for up to 4 weeks under a 50° C. condition. That is, thestability of excipient at 50° C. was tested.

As a result, as illustrated in Table 1 shown below, stearyl alcohol,phospholipon 90H, white wax, titanium dioxide, HPMC F4M, carbomer(carbopol 940), propylene glycol stearate, and aluminum starchoctenylsuccinate were found to be suitable, whereas Ceteareth-20 wasfound to be unsuitable.

TABLE 1 API: Day 0 2 weeks, 50° C. 4 weeks, 50° C. Excipient Appear-Recov- TRS. Appear- Recov- TRS. Appear- Recov- TRS. No. Excipient ratioance ery % % ance ery % % ance ery % % 1 Zileuton API — — 2.378 White101.47 3.311 White 99.12 2.711 powder 101.30 3.006 powder 98.23 2.493 2Stearyl 1:1 White 98.71 3.311 White 101.69 2.935 White 102.91 2.512alcohol powder powder 101.28 2.796 powder 98.77 2.567 3 Ceteareth-20 1:1White 100.34 2.394 White  99.99 3.445 White 97.59 3.632 powder power100.36 3.365 semisolid 96.39 3.549 4 Phospholipon 1:1 White 100.61 2.400White 101.50 2.820 White 97.26 2.557 90H powder powder 102.21 2.846powder 103.30 2.429 5 White wax 1:1 White 99.46 2.372 White 100.70 2.863White 98.77 2.459 powder powder 102.20 2.843 powder 97.31 2.381 6Titanium 1:1 White 100.59 2.344 White 102.55 2.803 White 99.14 2.507dioxide powder powder 102.86 2.839 powder 99.17 2.504 7 Hypromellose 1:1White 103.72 2.610 White 102.67 2.864 White 98.86 2.555 (HPMC) powderpowder 102.97 2.949 powder 100.86 2.580 8 Carbomer 1:1 White 99.34 3.326White 102.45 2.385 White 99.63 2.330 powder powder 103.39 3.067 powder97.65 2.403

In the solution stability test, suitability with solvents forsolubilizing zileuton (ethanol, PEG400, propylene glycol, hexyleneglycol, capryol 90, benzyl alcohol, water) was screened. For thispurpose, 10 mg of zileuton was dissolved in a 2 mL vehicle (solvent),and then changes of TRS % were observed in conditions of 25° C., 40° C.,and 70° C., respectively.

As a result, as illustrated in Tables 2 and 3 below, most of thesolvents were suitable to zileuton under the 25° C. condition, but wereunstable under the condition of 40° C. or 70° C. Table 2 shows theresult of stability test of the solution at 25° C., and Table 3 showsthe result of stability test of the solution at 40° C. or 70° C.

TABLE 2 TRS TRS (%) increase Solvents Temp. Day 0 Day 1 Day 3 Day 8 rate% Ethanol 25° C. 1.255 1.014 1.951 2.137 0.882 Polyethylene glycol 25°C. 1.427 1.491 2.897 4.065 2.638 (PEG-400) Propylene glycol 25° C. 1.4571.624 1.478 2.132 0.675 Hexylene glycol 25° C. 1.505 1.448 1.583 1.9800.475 Capryol 90 25° C. 1.623 1.444 1.478 2.201 0.578 Benzyl alcohol 25°C. 1.642 0.869 1.806 2.023 0.381 Water 25° C. 1.424 1.879 1.816 1.9650.541

TABLE 3 TRS TRS (%) increase Solvents Temp. Day 0 Day 1 Day 3 Day 8 rate% Ethanol 40° C. 1.255  1.778  2.375  4.704  3.449 Polyethylene glycol70° C. 1.427 43.725 94.366 97.142 95.715 (PEG-400) Propylene glycol 70°C. 1.457 26.382 82.228 96.325 94.868 Hexylene glycol 70° C. 1.505  8.06129.999 60.352 58.847 Capryol 90 70° C. 1.623 10.312 39.603 76.445 74.822Benzyl alcohol 70° C. 1.642 11.350 20.397 24.250 22.608 Water 70° C.1.424 35.171 80.268 91.994 90.570

Furthermore, the suitability of other various solvents for solubilizingzileuton was screened. For this purpose, solubility of zileuton in eachsolvent was observed in condition of 25° C.(RT), respectively.

As a result, as illustrated in Table 4 below, all of the solvents listedin Table 4 can be used for solubilize zileuton in manufacturing azileuton cream formulation. Especially, hydrophilics solvents such asalcohols may be preferably used.

TABLE 4 Zileuton Solubility Solvent at RT (% w/w) Triacetin 0.50-0.72Isopropyl Myristate <0.20 Propylene Glycol Dicaprylocaprate <0.20 MediumChain Triglycerides <0.21 Castor Oil <0.16 Olive Oil <0.07 Safflower Oil<0.07 Light Mineral Oil <0.04 Squalane <0.04 Dimethicone <0.06Cyclomethicone <0.06 Polyglyceryl-6 Dioleate <0.06 Polyglyceryl-10Mono/Dioleate/Caprol PGE-860 <0.16 Polyoxyl 40 Hydrogenated CastorOil/Kolliphor <0.37 RH40 Caprylocaproyl Polyoxyl-8 Glycerides/Labrasol10.81-11.42 Polyoxyethylene (20) Sorbitan 8.42-9.24 Monolaurate/Tween 20Diethylene Glycol Monoethyl Ether/Transcutol P 17.11-17.62

2. Composition Optimization Test

The components determined as being suitable in the aforementionedsuitability screen test were combined, and amount ratios of solvents andingredients stable under an acceleration condition (30° C., 40° C.) werescreened.

For this purpose, by mixing the ingredients other than solvents such aspropylene glycol and water and the like under 70° C. according to theamount ratios disclosed in Table 5, and then adding and mixing theremaining solvents while stirring for 30 minutes under a condition of70° C. and 800 rpm or less. The mixed composition was stirred at 100 rpmor less, and cooled to room temperature. That is, Table 5 below showsresults of the test regarding optimized ingredient ratios.

TABLE 5 samples Ingredients (wt %) 2-1 2-2 2-3 2-4 2-5 2-6 2-7 2-8 2-92-10 2-11 2-12 Zileuton — — — — — — — — — — — — Water — — — 57 16 — —10~31 White petrolatum 43 38~48 24 20 43 38 18~31 White wax 6 6 6 6 5 77 5~9 Propylene 24 32~42 10 55 47 52 25~55 glycol(PG) Stearyl Alcohol 16— — — — — — — — — — — Propylene glycol — — — — — — — — — — — — stearatePhospholipon 90H — 3 3 3 4 3 3 4 4 4 4 4 Ethanol — — — — — — — — — —10.5 10.5 Aluminum 11 11 11 — — — — — — — — — starch octenyl- succinateAppearance Little Cream Cream Liquid Oint- Oily Very Cream Cream CreamCream Cream dry ment cream oily cream cream Approximate — — — — <1.3%<3% — <0.1% <0.3% <0.7% <2.3% <2.5% solubility ( % )

As shown in Table 5, it was found the composition ratios of 2-2, 2-3,2-8, 2-9, 2-10, 2-11, and 2-12 provide suitable cream type formulation.From the aforementioned result, it was confirmed that the amount ratiosof water, white petrolatum, white wax, and propylene glycol havecritical significance in the aforementioned ranges.

3. Optimization Test of Cream Processing Method

A condition that satisfies the physical and chemical stability with atarget drug concentration (1%) was screened based on the amount ratiosdiscovered in the composition optimization test. For this purpose, theingredients other than the solvents such as propylene glycol and waterwere mixed according to the amount ratios disclosed in Table 6 under 70°C., and then the remaining solvents were mixed at 70° C. and 800 rpm orless. The mixed cream composition was stirred at or less than 100 rpmand cooled to room temperature.

Physical stabilities were measured by checking with the naked eyewhether or not phase separation occurred after leaving a sample for acertain period of time at a condition of 25° C., 30° C. or 40° C. Thisis a test to discover an optimized processing method for obtaining acream formulation.

TABLE 6 Ingredients samples (wt %) 3-1 3-2 3-3 3-4 3-5 3-6 3-7 3-8 3-93-10 3-11 3-12 Zileuton — — — — — — — — — — — — Water 10~12 17~21 White18~20 17~21 petrolatum White wax 6 6 6 6 6 6 6 6 6 6 6 6 Propylene 51 5153 55 55 51 54 51 48 52 54 56 glycol(PG) Phospholipon 4 4 4 4 4 5 5 4 44 4 4 90H Ethanol 4.5~9   — — — — Stirring 20 20 20 20 20 20 20 20 20 2020 20 time(min) Shape Cream Cream Cream Cream Cream Cream Cream CreamCream Cream Cream Cream Approximate <2.1% <1.8% <2.7% <2.7% <2.3% <2.1%<2.1% <2.1% <1.3% <1.5% <1.7% <2.1% solubility (%) Physical Bad Bad BadBad Bad Bad Bad Bad Good Good Good Good stability at 25° C.

As disclosed in Table 6, it was found that the formulation conditions of3-9, 3-10, 3-11, and 3-12 provide excellent physical stability at 25° C.so as to dissolve active pharmaceutical ingredient 1% or more.Furthermore, the aforementioned cream formulations do not compriseethanol, and this is because ethanol deteriorates physical stability.

Next, as a test for optimizing the processing method for cream typeformulation, in order to find out whether the stirring time affects theshape of the cream, stirring times of 5 minutes, 30 minutes and 60minutes were applied and then observed. As a result, as illustrated inTable 7 below, the stirring time of 30 minutes was confirmed as theoptimized condition.

TABLE 7 samples Ingredients(wt %) 3-13 3-14 3-15 Zileuton — — — Water10~26 White petrolatum 17~29 White wax 6 6 6 Propylene glycol(PG) 35~56Phospholipon 90H 4 4 4 Ethanol 6.5 — — Total sum of ingredients 100 100 100  Stirring time(min) 5 30  60  Appearance Liquid like Cream CreamCream Approximate solubility (%) — 0.3% <1.3% Physical stability at 30°C. Good

4. Optimization Test of Solvent

It was determined that using an excessive amount of propylene glycol toadjust to 1% of drug concentration has a negative effect on the physicalstability of the composition, and thus a composition ratio of highstability with a lower drug concentration was sought for.

Therefore, in order to improve the physical stability of the compositionafter increasing the scale of sample, a cream type formulation with areduced loading amount of drugs was prepared. For this purpose,formulation 3-14 was selected, and based on this cream type composition,the amount of drug loading was raised up to 1%, and then a solventcapable of obtaining chemical stability while retaining physicalstability was screened.

The aforementioned screening test was conducted by mixing ingredientsother than the solvents such as propylene glycol, water and the like at70° C. in accordance with the composition ratio disclosed in Table 8below, and then mixing the remaining solvents according to thecomposition ratio of Table 8 under a stirring condition of 800 rpm orless for 30 minutes at 70° C. The mixed composition was stirred at orless than 100 rpm and cooled to room temperature.

The chemical stability was evaluated by measuring changes in TRS %through LC analysis after leaving a sample for a certain period of timeunder a temperature of 25° C. or 40° C.

TABLE 8 samples N-methyl pyrrolidone Ingredients (NMP) Capyrol 90 Benzylalcohol (wt %) 4-1 4-2 4-3 4-4 4-5 4-6 4-7 4-8 4-9 4-10 Zileuton — — — —— — — — — — Water 24~26 20~24 37~41 White 29~33 29~33 38~42 petrolatumWhite wax 6 6 6 12~24 6 Propylene 24 24 24 — — — — — — — glycol(PG)Stearyl — — — — — — 4 — — — Alcohol Propylene — — — — — — — 4 — — glycolstearate Phospholipon 4 4 4 4 4 4 — — 4 4 90H Ethanol — — — — — — — — —— Aluminum — — — — — — — — — — starch octenyl- succinate N-methyl 10 108 — — — — — — — pyrrolidone (NMP) Benzyl — — — — — — 14 14 10 10 alcoholCapyrol 90 — — — 28 28 28 — — — — Dimethyl — — — — — — — — — — sulfoxideStirring time 30 30 30 30 30 30 30 30 30 30 (min) Appearance LiquidLiquid Cream Cream Cream Cream Cream Cream Cream Cream Approximate 1% 1%0.8% 0.8% 0.8% 0.8% solubility (%) Physical Bad stability at 30° C.Physical Bad Bad Bad Bad Bad Bad Bad stability at 40° C.

As illustrated in Table 8, NMP and benzyl alcohol were screened assolvents, but both showed bad physical stability with phase separation.Furthermore, capryol 90 was screened as a solvent, but all showed badphysical stability due to phase separation.

Consequently, the processing method was changed from 3-14 condition, andan amount of drug loading was increased to 1%, and thereby a cream typecomposition that satisfies both the target drug concentration andphysical stability was discovered. However, regarding the chemicalstability, a significant TRS % change was observed after 3 weeks under atemperature condition of 40° C.

5. Test for Confirming an Improvement in Stability of the Composition byUse of a pH Buffer Solution as Solvent

In order to confirm the effect of a pH buffer solution on the chemicalstability of the composition, the compositions were prepared by mixingingredients other than the solvents at 70° C. in accordance with thecomposition ratios disclosed in Table 9 below, and then mixing theremaining solvents according to the composition ratios of Table 9 undera stirring condition of 800 rpm or less for 30 minutes at 70° C. Themixed compositions were stirred at or less than 100 rpm and cooled toroom temperature.

The chemical stability was evaluated by measuring changes in pH,quantitative analysis of degradants of zileuton by HPLC, and TRS %through LC analysis after leaving a sample for a certain period of timeunder the storage condition of a room temperature of 25° C.(RT) and 40°C.

TABLE 9 samples Ingredients(wt %) 5-1 5-2 Zileuton 1.0 1.0 pH buffersolution* — 32 Water 32 — White petrolatum 36-40 White wax 4-8 Propyleneglycol 16-22 Phospholipon 90H 2-6 Propylparaben 0.005-0.04  Total sum ofingredients 100 Appearance Cream Cream *pH buffer solution at pH 4, 5and 6 with various concentration

As illustrated in FIG. 1, pH changes of the pH buffered creams at pH 4and pH 5 were lowered in comparison with the non-buffered cream underaccelerated condition (40° C.). It is noted that the pH buffered creamwith a pH buffer solution of pH 4 can be controlled between pH 4.5 to 5at which the generation of hydrolytic degradants of zileuton, i.e.impurities, are mostly suppressed, even at 40° C. when 10 mM to 100 mMpH buffer solutions were used, while the pH of the non-buffered controlsample was changed to ˜pH 7.

Furthermore, as illustrated in Table 10 below, the changes of TRS (totalrelated substances) of the non-buffered control composition wasincreased up to 9.96% at 40° C. for 2 months, while the changes of TRSof the buffered composition with 25 mM pH buffer solution of pH 4 wasonly increased up to 2.74% in the same condition.

TABLE 10 40° C./3 Weeks 40° C./2 months *Initial non- pH 4 pH 4 non- pH4 pH 4 non-buffered buffered 10 mM 25 mM buffered 10 mM 25 mM Totalrelated 0.23% 2.18% 1.60% 1.52% 10.19% 3.29% 2.97% substance % Change ofTRS — 1.95% 1.37% 1.29% 9.96% 3.06% 2.74% *Initial: day of formulation

6. Optimization Test of the Amount of Ingredients

Furthermore, as Illustrated in Table 11 Below, a Test for Determining aCritical Amount range of ingredients was conducted after selecting aningredients and an approximate composition ratio that could satisfycriteria as a topical cream.

TABLE 11 samples Ingredients (wt %) Criteria 6-1 6-2 Zileuton — — —Water 30~40 32 32 White petrolatum 36~40 33 38 White wax 4~8 6 6Propylene glycol 16~22 25 19 Phospholipon 90H 2~6 4 4 Appearance CreamCream Cream Physical stability for 4 Stable Phase separation Stableweeks under 40° C. observed at the 4^(th) week

As shown in Table 11, as in the formulation 6-1, when the amount ratioof white petrolatum and propylene glycol are 33 and 25, respectively, itwas observed by naked eyes that the physical stability was deteriorateddue to separation of oil phase and water phase starting from the 4^(th)week.

Table 12 below shows candidate formulations according to the testresult.

TABLE 12 Candidate formulations samples Ingredients (wt %) 6-3 6-4Zileuton 1 1 Water 32 26 White petrolatum 32 29 White wax 6 6 Propyleneglycol (PG) 25 34 Phospholipon 90H 4 4 Total sum of ingredients 100 100Stirring time (min) 30 30 Appearance Cream Cream Approximate solubility(%) 1% 1% Physical stability at 40° C. Good Good Chemical stability at40° C. 6.65% of changes 9.79% of changes (TRS %) for 3 weeks for 3 weeks

As shown in Table 13, the formulations 6-3 and 6-4 satisfied conditionsrequired for development up to clinical phase 2. Further, in order todevelop the final formulation, an experiment for developing a newcomposition ratio and processing method was additionally conducted whilemeasuring the chemical stability under 30° C. acceleration condition of062-1.

Accordingly, as shown in Table 13 below, the amount of propylene glycolwas optimized, and propylene paraben was added as a preservative. As aresult, the cream formulations 6-5 and 6-6 of the composition ratios ofTable 13 satisfied chemical/physical stabilities under 30° C.acceleration condition.

TABLE 13 samples Ingredients(wt %) 6-5 6-6 zileuton 0.1 1 Water 30-3430-34 White Petrolatum 36-40 36-40 White wax 4-8 4-8 Propylene glycol16-22 16-22 Phospholipon 90H 2-6 2-6 Preservative (Propylparaben)0.05-0.4  0.05-0.4  Total sum of ingredients 100 100 Stirring time (min)30 30 Appearance Cream Cream Physical stability Good for 4 weeks Goodfor 4 weeks Chemical stability (TRS %) 1.4% of changes for 1.4% ofchanges for 4 weeks at 30° C. 4 weeks at 30° C.

On the other hand, it was confirmed that the cream formulations of thecomparative examples having composition ratios of Table 14 below do notexhibit good chemical/physical stabilities under 30° C. accelerationcondition in the target drug concentration range which can provideefficacious pharmaceutical activity.

TABLE 14 samples Ingredients(wt %) 6-7 6-8 zileuton 0.7 0.7 Water 24 22White Petrolatum 25 27 White wax 3 3 Propylene glycol 46 46 Phospholipon90H 1 1 Preservative (Propylparaben) 0.02 0.02 Stirring time (min) 30 30Appearance Cream Cream Physical stability Phase separation Phaseseparation observed at 30° C. observed at 30° C.Experiment 1 Evidence of Concept Research Using Formulation 089

An in vivo test was conducted to identify whether or not the formulation6-6 of the above example shows medical effects when administered to skinin a generally used mouse atopic dermatitis animal model. For thispurpose, BALB/c mice were divided into 4 groups, each consisting of 10mice. The groups were divided into a vehicle (acetone), positive controlgroup (dexamethasone 0.05 mg/ear; Dex), and zileuton groups (Formulation5-6). And the zileuton groups were divided into cases where acetone (1mg/ear) and the cream formulation of 6-6 (0.2 mg/ear and 0.02 mg/ear)were used, respectively.

As illustrated in FIG. 2, on day 0 and day 1, 25 μl of 0.5% DNFB(dinitrofluorobenzene) solution was applied once on ears of all groupsto cause inflammation. Then, on day 5, 20 μl of 0.3% DNFB was appliedonce again to cause inflammation in the ears. Then, dexamethasone andzileuton dissolved in acetone vehicle were applied to the ears at 1hour, 6 hours, and 23 hours after the stimulation on day 5. Thicknessesof the ears were measured at 24 hours after the stimulation on day 5,and then ear tissues were taken for histopathological observation.

As illustrated in FIG. 3, in the group where the formulation 6-6 wasused, zileuton (API: Active Pharmaceutical Ingredient) was effectivelydelivered to the skin, thereby effectively inhibiting increase of earthickness caused by inflammation and edema compared to groups whereacetone was used.

Through histopathological observation, excellent anti-inflammationfunction of zileuton was observed.

TABLE 15 Test model BALB/c mouse, female 6~8 weeks of age (17~20 g)Dosing Drug Concentration mL or API Regimen Group (formulation) No.Route (mg/mL or %) mg/ear mg/ear (Time) 1 Vehicle 10 Topical N/A 0.02 mLN/A 1, 6, 23 (Acetone) 2 Dex 10 Topical 2.5 mg/mL 0.02 mL 0.05 1, 6, 23(Acetone) 3 Zileuton 10 Topical 50 mg/mL 0.02 mL 1 1, 6, 23 (Acetone) 4Zileuton 10 Topical 1 wt % 20 mg 0.2 1, 6, 23 (formulation 5-6) 5zileuton 10 Topical 0.1 wt % 20 mg 0.02 1, 6, 23 (formulation 5-6 exceptof zileuton 0.1 wt %) Measure Weight Ear thickness Histopathologicalobservation (edema, inflammation, crust formation)Experiment 2 PK Research Test Using Minipigs

It is necessary to find out from the Experiment 1 whether or not medicaleffects by absorption to an entirety of body are included. In order toidentify the skin absorption and systemic exposure level of the finalformulation, minipigs having skin structures most similar to those ofhumans may be used. Therefore, skin PK was observed in minipigs havingthe most similar skin structures as humans using the formulation 6-6which is for the clinical formulation derived in the process ofdeveloping the cream formulation.

For this purpose, 20 mg of zileuton (API: Active PharmaceuticalIngredient) was applied to 100 cm² of skin using the formulation 6-6.After applying the formulation 6-6, concentration of zileuton in plasmawas measured 0.25, 0.5, 1, 1.5, 2, 3, 4, 6, and 8 hours afterwards.After applying the formulation 6-6, zileuton concentration in the skinwas measured 1 and 8 hours afterwards. For the measurement of zileutonconcentration in the skin, the skin was separated into surface,subcutaneous, dermas, and epidermis layers. The subcutaneous layer wasseparated using the tape stripping method, and the measurements ofzileuton concentrations were conducted for different depths of strip 1,strip 2-5, strip 6-12, and strip 13-24. Test conditions are asillustrated in Tables 16 and 17.

TABLE 16 Treatment Concen- Number of tration (mg Type Bamma pigs TestDose API/g of specimen Group (#/Gender) drug (mg/cm²) cream) VehicleRoute obtained 1 6 females Zileuton 0.2(API) 10 6-6 Topical Plasma,(Q301) Formulation administration skin Test article storage: Comments:Animals of body weights of 15~20 kg were used, Desiccated at 4° C. andadministration area was 10 cm × 10 cm Overnight Fast of Animals: None

TABLE 17 Adminis- Animal Sampling and administration schedule (time)Group tration route ID −0.5 0 0.25 0.5 1 1.5 2 3 4 6 8 1 Topical 1~3 P DP 0 P + S — — — — — — 4~6 P D — P — P P P P P P + S P: Collect blood andseparate plasma D: Administer drug to animals at determined time P + S:Collect blood and skin, and separate plasma from blood. Extract corneumlayer before obtaining skin sample. Separate dermas and epidermis layersfrom skin. Anticoagulant: K2-EDTA

As illustrated in FIGS. 4 and 5, the formulation 6-6 showed effectiveskin absorption pattern, whereas it did not show any systemic exposurepattern in the minipigs. It can be seen that the cream formulation usingpropylene glycol effectively induces skin absorption, and minimizes bodyabsorption, and is thus suitable as a future candidate formulation ofzileuton for skin administration.

While this invention has been described in connection with what ispresently considered to be preferable embodiments, it is to beunderstood that the present invention is intended to cover variouschanges, modifications and equivalent arrangements. It is obvious thatthe present invention may be practiced by appropriate modifications ofthe aforementioned embodiments. Therefore, the aforementionedembodiments do not limit the scope of the present invention which isdetermined by the claims.

INDUSTRIAL APPLICABILITY

The present invention is for providing a zileuton cream formulation fortopical treatment of skin disease caused by leukotriene, and is anindustrially applicable invention.

We claim:
 1. A topical anti-inflammatory pharmaceutical composition,comprising zileuton, an aqueous solvent, an organic solvent, athickening agent, and an emulsifier, wherein the composition compriseszileuton of about 0.05 to about 2 weight %; pH buffer solution of about30 to about 34 weight %; white petrolatum of about 36 to about 40 weight%; white wax of about 4 to about 8 weight %; propylene glycol of about16 to about 22 weight %; phospholipid of about 2 to about 6 weight %;and preservative of about 0.005 to about 0.04 weight %.
 2. Thecomposition according to claim 1, wherein the pH buffer solution has apH of about 4 to about
 5. 3. The composition according to claim 2,wherein the pH buffer solution is citrate buffer, acetate buffer orphthalate buffer.
 4. The composition according to claim 1, wherein thephospholipid is phospholipon 90H.
 5. The composition according to claim1, wherein the preservative is at least one selected from a groupconsisting of methylparaben, ethylparaben, propylparaben,isobutylparaben, butyl paraben, 2-phenoxy ethanol, and 4-hydroxybenzoicacid.
 6. The composition according to claim 1, wherein the compositioncomprises zileuton of about 0.1 to about 1 weight %; water or pH buffersolution of about 31 to about 33 weight %; white petrolatum of about 37to about 39 weight %; white wax of about 5 to about 7 weight %;propylene glycol of about 18 to about 20 weight %; phospholipid of about3 to about 5 weight %; and preservative of about 0.01 to about 0.03weight %.
 7. The composition according to claim 1, wherein thecomposition is formulated for topical application to human skin.
 8. Thecomposition according to claim 1, wherein zileuton is racemic zileuton.9. A method for relieving or treating atopic dermatitis, acne,urticaria, psoriasis, eczema, a bullous skin disease, collagenoses,Sjogren-Larsson syndrome, or acne in skin lesions of mastocytosis,wherein said method comprises administering, to a subject in need ofsuch relief or treatment, a composition of claim
 1. 10. The methodaccording to 9, wherein the composition is applied topically to humanskin.
 11. The composition according to claim 1, wherein the compositionis a cream.